A biofilm is a community of microorganisms attached to a surface, surrounded by an extracellular polymeric substance (EPS) made of polysaccharides, DNA and proteins, able to protect microbes from environmental assaults. Bacterial attachment to surfaces and consequent biofilm formation are known phenomena in diverse environments such as medical and food industry devices.
Bacteria growing as a biofilm remain a significant challenge as they tend to be more tolerant to antimicrobial treatments.
Biofilms are an important cause of infections associated with biomaterials and urinary tract infections (UTIs) and are an important cause of nosocomial infections with significant morbidity, mortality and additional hospital costs.
Intact host defence systems usually eliminate transient bacterial contamination or colonization, but the presence of foreign bodies, such as catheters and implants, lowers the threshold of bacterial sustainable infection and generates local immunosuppression allowing biofilm formation on foreign surfaces in the human body, that tends to be recalcitrant to standard antimicrobial treatment. The biofilm formation can, in fact, immediately start once a biomedical device has been placed in its body niche. As frequent replacement of the prosthesis due to bacterial infection is uncomfortable, costly, time consuming and may lead to damage of the cellular tissue of patients, alternative approaches to catheter infection treatments must be found. Approximately 80% of nosocomial UTIs are associated with the use of indwelling urinary catheters and UTIs are responsible for 40-60% of all hospital-acquired infections. Bacteria found adhering to the intraluminal surfaces of catheters are the principal source and cause of bloodstream infections. Approximately five million central venous catheters are inserted per year, and of these 3-8% lead to bloodstream infection. The attributable mortality of these bloodstream infections is 12-25%. In contrast, the mortality rate of catheter-associated UTI is less than 5%. Escherichia coli is one of the most prevalent pathogens in UTIs such as prostatitis in adult men and cystitis in women.
Biofilm infection and its correlated diseases can be limited by preventing microbial adhesion to tissues and/or medical device surfaces but also an effective therapy to eliminate an existing biofilm is desirable.
Biosurfactants, amphiphilic metabolites produced by a wide group of bacteria from various biochemical building blocks, can be a useful approach to challenge bacteria growing as a biofilm. Microbial biosurfactants include a wide variety of surface-active compounds such as glycolipids, lipopeptides, polysaccharide-protein complexes, phospholipids, fatty acids and neutral lipids. Among the many classes of biosurfactants, lipopeptides (surfactin, iturin and fengycin classes) are particularly interesting because of high surface activity and antibiotic potential.
surfactin is a mixture of cyclic lipopeptides built from variants of a heptapeptide and a β-hydroxyl fatty acid with chain length of 13-15 carbon atoms. It is produced by various Bacillus strains. A lactone bridge between the β-hydroxyl function of the acid and the carboxy-terminal function of the peptide confers a cyclic structure to this molecule. A natural diversity occurs, giving rise to homologues differing from each other by the acyl chain length (13 to 15 atoms of carbon) and further isoforms are characterized by differences in the amino acid sequence.
fengycin family consists of a β-hydroxy fatty acid connected to the N-terminus of a decapeptide, including four D-amino acid residues and the rare amino acid L-ornithine. The C-terminal residue of the peptide moiety is linked to the tyrosine residue at position 3, forming the branching point of the acylpeptide and the eight-membered cyclic lactone. fengycin consists of two isoforms, fengycin A and B, which differ in their amino acid sequences. fengycin A presents Ala at position 6, fengycin B presents Val at the same position. The length of the β-hydroxy fatty acid tail is variable (from C14 to C18) and links the amino group of its N-terminal amino acid Glu. Different homologous compounds for each lipopeptide family are thus usually coproduced by the same bacterial strain.
Biosurfactants have advantages over their synthetic chemical counterparts because of their biodegradability and reduced toxicity, availability from cheap raw materials, biocompatibility and the effectiveness at extreme temperature, pH and salinity. Biosurfactants have found possible applications in biomedical fields.
Organisms within a biofilm are difficult to eradicate by conventional antimicrobial therapy and can cause indolent infections. Resistance of bacterial infections to antibiotics is increasing worldwide and some older antimicrobial agents are no longer recommended because of high levels of resistance.
Antibiotics and biocides may kill the free-living microorganisms (planktonic), but are not fully effective in killing organisms in a biofilm, leaving viable cells on surfaces. EPS confers protection from cellular immunity, by preventing phagocytosis, and from antibiotics. In addition, biofilms show increased resistance to antibiotics by decreasing antibiotic penetration through EPS, upregulating multi-drug efflux pump expression and expressing periplasmic glucans that directly bind to and sequester antibiotics.
Bacteria in deep biofilm layers can be less metabolically active, compared with their planktonic counterparts and, thus, appear to be more resistant to antibiotics. Bacteria living in a biofilm can create self-generated diversity that insures the survival of microorganisms sharing enzymes and proteins produced by other community members.
Several attempts have been made to avoid biofilm formation by incorporation of biocides into surface materials or by coating surfaces with biocides, especially in the medical field. It has been suggested that strategies for antimicrobial use in UTIs should be based on more potent antimicrobial agents, not necessarily broader-spectrum agents, and appropriate dosage and duration of antibiotic treatment.